DSN normalisation

[Abdul K Sesay]

We are trying to produce a cDNA library for RNA-Seq from 100ng of total RNA using the DSN normalisation protocol from your website. Earlier in the week, we were told by Illumina to follow the mRNA Sequencing Sample Preparation Guide, starting at 'Fragment the RNA', up to ligation of the adaptors, and then starting the DSN normalisation protocol. However, there doesn't appear to be any size selection if we do it this way, can this be right?

Also, one of the pdfs on Illumina website suggests that we should enrich the library by PCR before starting DSN normalisation:-

http://www.illumina.com/documents/seminars/presentations/2010-06_sq_03_lakdawalla_transcriptome_sequencing.pdf

Could you help us by clarifying which steps of the mRNA-Seq protocol we should omit when producing a library from 100ng of total RNA?

Many thanks in advance,

Abdul